We propose to use the technique of direct scanning gel chromatography to investigate the interaction of enzymes and substrates, and enzyme protein components with one another. We have built a direct scanning chromatography to use for this purpose. We will modify it to include automated operation and an interactive system of data reduction. A PDP 1104 minicomputer will be used as the controller and data processer. We will examine the association-dissociation of hexokinase as it is affected by the presence of substrates glucose and of ATP or their analogs CrATP and lyxose. Both direct scanning and active enzyme chromatography will be used to characterize the association behavior of the enzyme chromatography will be used to characterize the association behavior of the enzyme and differential chromatography will be used to measure conformation changes induced by substrates or analogs. We will then undertake an examination of the interaction of nitrogenase components with one another as this interaction is influenced by the presence of substrates ATP and dithionite. We will measure the association to complexes by use of active enzyme chromatography following the oxidation of dithionite. Comparisons will be made between combinations of the nitrogenase components of different microorganisms and mutants which are defective in nitrogen reduction. We will investigate the interaction of cytochrome oxidase with cytochrome c by direct scanning chromatography. An optical transition, possibly associated with a conformational change in the oxidase, will be followed in both broad zone and differential chromatography to determine the nature of binding of cytochrome c to the oxidase, (i.e., the relative tightness of binding in different combinations of oxidized and reduced components).